The HIV-1 envelope (env) protein exists as an oligomeric complex on the surface of virions and infected cells. Work from our lab and several others has indicated that env oligomeric structure has important implications for understanding the humoral immune response, and may well be important for eliciting broadly cross-reactive, neutralizing antibodies. However, it is also clear that much vaccine work that has concentrated on T-cell line adapted HIV-1 strains, including some of our own, has probably been misguided. Rather, env proteins derived from primary virus isolates should be studied in their place. Recent breakthroughs in the chemokine receptor field have further served to highlight differences between lab adapted and primary virus isolates. Therefore, we have shifted our focus to primary virus isolates, particularly the dual-tropic virus strain 89.6. We have developed techniques to obtain milligram quantities of purified, soluble, monomeric and oligomeric forms of primary env proteins, and propose and highly collaborative project designed to test the efficacy of both DNA and subunit vaccination strategies in a rigorous manner. We will generate, characterize, and produce in milligram quantities primary HIV-1 env proteins. These proteins will then be used by our colleagues in a series of immunization studies. Of these, perhaps the most important are those that will attempt to confer immunity in rhesus macaques to a lethal challenge with SHIV 89.6P. We will then assist in the characterization of the humoral immune response during vaccination and after virus challenge both for this and other collaborative projects. Furthermore, we will investigate the antigenic structure of primary virus env proteins by producing MABs to oligomeric 89.6, and will investigate antibody neutralizing mechanisms in light of our rapidly increasing knowledge of env-chemokine receptor interactions.